一种新型免疫层析技术在临床乳腺癌Her-2蛋白定量检测中的应用

建立一种靶点蛋白质快速定量检测方法。在原有侧向流动免疫层析技术的基础上,通过优化层析材料和纳米微球的均一性、改进检测区的检测方法,经逐点扫描技术,建立标准浓度曲线,以达到对临床靶点蛋白质的定量检测。以乳腺癌组织中的Her2表达为例,通过对已知浓度样品的检测,验证本技术方法的准确度大于96%。另外,以蛋白质免疫印迹作为组织中特定蛋白质检测金标准,分析临床肿瘤组织中Her2蛋白的含量,其准确率也达到95.5%,而免疫组织化学方法检测准确率仅为69.58%。新型免疫层析法检测结果与靶向治疗患者的愈后密切相关(P<0.01)。改进后的新型免疫层析方法能够准确地对临床靶点蛋白质进行定量检测,而且结合侧向流动技术的简单、快速和易用性,这种新型检测方法可以广泛应用于临床组织标本、血液标本和体液标本中靶点蛋白质的临场定量检测,在一定程度上可以替代免疫组化技术。     

Soluble expression and purification of Brucella cell surface protein (BCSP31) of Brucella melitensis and preparation of anti-BCSP31 monoclonal antibodies

Brucella cell surface protein (BCSP31) is potentially useful for diagnosing brucellosis. We aimed to establish a monoclonal antibody (MAb) against Brucella melitensis BCSP31 and to investigate its distribution in diagnosis. Soluble recombinant BCSP31 was successfully expressed and purified. Two MAbs (1F1 and 1E5) against B. melitensis BCSP31, effective in detecting both recombinant and cellular proteins, were obtained and characterized. The MAbs did not react with Escherichia coli, Staphylococcus aureus, Bacillus subtilis, Mycobacterium tuberculosis, or Bacillus aeruginosus, but strongly reacted with BCSP31 and B. melitensis by ELISA and Western blot analysis. We also tested different Brucella species and brucellosis using the prepared anti-BCSP31 MAbs. BCSP31 and anti-BCSP31 MAbs may play important roles in future research in diagnosing brucellosis.     

eEF-2 Phosphorylation Down-Regulates P-Glycoprotein Over-Expression in Rat Brain Microvessel Endothelial Cells

ObjectiveWe investigated whether glutamate, NMDA receptors, and eukaryote elongation factor-2 kinase (eEF-2K)/eEF-2 regulate P-glycoprotein expression, and the effects of the eEF-2K inhibitor NH125 on the expression of P-glycoprotein in rat brain microvessel endothelial cells (RBMECs).MethodsCortex was obtained from newborn Wistar rat brains. After surface vessels and meninges were removed, the pellet containing microvessels was resuspended and incubated at 37°C in culture medium. Cell viability was assessed by the MTT assay. RBMECs were identified by immunohistochemistry with anti-vWF. P-glycoprotein, phospho-eEF-2, and eEF-2 expression were determined by western blot analysis. Mdr1a gene expression was analyzed by RT-PCR.ResultsMdr1a mRNA, P-glycoprotein and phospho-eEF-2 expression increased in L-glutamate stimulated RBMECs. P-glycoprotein and phospho-eEF-2 expression were down-regulated after NH125 treatment in L-glutamate stimulated RBMECs.ConclusionseEF-2K/eEF-2 should have played an important role in the regulation of P-glycoprotein expression in RBMECs. eEF-2K inhibitor NH125 could serve as an efficacious anti-multidrug resistant agent.     

厚壳贻贝whirlin类贝壳基质蛋白的重组表达与功能分析

贝壳历来是生物工程和材料学研究的重要对象。贝壳中的贝壳基质蛋白质在贝壳的形成与发育过程中具有重要的调控作用。Whirlin类蛋白质(Whirlin-like protein,WLP)是一种从厚壳贻贝(Mytilus coruscus)中鉴定的新型贝壳基质蛋白质。序列分析结果显示,该蛋白质含有PDZ(postsynaptic density/Discs large/Zonula occludens)结构域,而该结构域对贝壳生物矿化的影响目前尚无报道。为深入了解WLP在贝壳形成中对碳酸钙晶体的影响,在序列分析基础上,采用密码子优化结合原核重组表达,获得其重组表达产物后,开展了重组WLP对碳酸钙晶体形貌及晶型的影响研究,结晶速度抑制以及碳酸钙晶体结合分析。分析结果表明,重组WLP能诱导文石型碳酸钙晶体的形貌和方解石型碳酸钙晶体的晶型发生改变;同时重组WLP对碳酸钙晶体具有结合作用,且能抑制碳酸钙晶体的结晶速度。上述结果表明,WLP对贝壳的形成及发育具有重要影响,并可能在贝壳肌棱柱层的形成中发挥了重要作用。     

典型矿区植被覆盖度时空分布特征及影响因素

植被状况可以直接或间接地反映采煤对生态环境的影响。以长河井工煤矿、离柳井工煤矿、平朔露天煤矿3个典型矿区为研究区域。以Landsat数据为数据源，基于地形调节植被指数的像元二分模型提取植被覆盖度；采用趋势分析、线性回归斜率、稳定性分析方法，分析了3个典型矿区2001-2016年植被覆盖度的时空变化特征；运用"以时间换空间"的方法，采用相关分析方法对植被覆盖度变化的自然影响因素进行了分析。结果表明：（1）近16年3个典型矿区植被覆盖度呈增加趋势，长河、离柳、平朔矿区的增长速率分别为0.09%/10 a、0.10%/10 a、0.08%/10 a（P > 0.05）。（2）空间上，长河、离柳、平朔矿区植被覆盖度变化不明显比例分别占到66.63%，59.90%，62.25%，呈增加趋势的比例仅分别占28.14%、32.55%、27.81%，而呈减少趋势的比例分别占到5.23%、7.55%、9.94%。长河矿区明显改善的区域位于自然植被和耕作区的北部和东北部，离柳矿区明显改善的区域位于以低植被覆盖度为主的北部，平朔矿区明显改善的区域位于复垦的中西部。（3）不区分植被类型时，3个矿区的植被覆盖度变化与高程、高程与温度的交互作用表现出显著相关性（P < 0.01），与各自然因素的相关性总体表现为长河 > 离柳 > 平朔矿区；区分植被类型时，草地与坡度的相关性不显著（P > 0.05），与降雨量、高程存在显著正相关（P < 0.05）；灌木林与温度相关性不显著，与高程和降雨量的交互作用存在显著正相关；旱地与高程、高程与温度的交互作用存在显著相关性；疏林地与坡向、降雨量与坡向坡度的交互作用均没有表现出相关性；有林地与高程降雨量的交互作用表现出显著正相关性。探讨不同植被类型对自然因素的响应，可为矿区植被结构的选择，矿区复垦提供参考依据。